1. Field of the Invention
The present invention relates to the field of haemostasis and in particular to the aspect of thrombosis. More particularly the invention is directed at a method for screening and diagnosis of thrombophilia, especially hereditary thrombophilia. The method according to the invention can then be used for determining the risk for thrombosis in individuals.
2. Description of the Related Art
Thrombosis is a principal human affliction, killing hundreds of thousands and debilitating millions each year by myocardial infarction, pulmonary embolism, or stroke. Risk factors include both hereditary and acquired conditions. Generally, a tendency towards thrombosis could arise from hyperactive coagulation pathways, hypoactive anticoagulant mechanisms, or hypoactive fibrinolysis. Mutations in genes that encode proteins in these pathways play an important role in the predisposition to thrombosis. Variant alleles of the genes encoding protein C, protein S, antithrombin, and fibrinogen have been shown to be relatively strong, but uncommon risk factors for thrombosis. Genetic analysis of these genes showed a large heterogeneity of mutations. More recently, a poor anticoagulant response of plasma to activated protein C (APC) due to the presence of a mutant factor V molecule (factor V R506Q, factor V Leiden) was discovered and is as yet the most common hereditary risk factor for thrombosis known. Recently some support was obtained for the hypothesis that the clustering of thrombosis in families is due to epistatic effects. Studies in selected families with venous thrombosis indicated that the presence of mutations in two genes may increase the penetrance of the thrombotic disease.
The discovery of genetic risk factors for thrombosis came after the identification of families in whom the thrombophilia segregated with an abnormal result in a plasma test (protein C, protein S, antithrombin, and APC resistance). However, despite the ever growing insight into the processes of coagulation and fibrinolysis, the underlying cause of many inherited thrombotic events remained unsolved.
The serine protease thrombin, formed by cleavage of human prothrombin, exerts a central action to the processes of thrombosis and haemostasis. The thrombin molecule plays a role in the final stage of blood coagulation: the formation of an insoluble fibrin clot. The hithereto known congenital disorders of prothrombin are rare and involve either reduced synthesis of an (ab)normal molecule (referred to as hypoprothrombinemia or type I prothrombin deficiency) or the normal synthesis of a dysfunctional molecule (referred to as dysprothrombinemia or type II prothrombin deficiency). Homozygous patients with hypoprothrombinemia generally have plasma prothrombin levels between 1% and 25% of normal values, conditions that are usually associated with a clinical bleeding tendency. Heterozygous patients, in contrast, have prothrombin levels of about 50% and at worst very mild bleeding problems. Patients with dysprothrombinemia have only 2% to 50% of the clotting activity of normal prothrombin; in these patients the severity of the bleedings correlates fairly well with the amount of prothrombin activity in plasma. A number of dysprothrombinemias have been further characterized by amino acid sequence analysis of the isolated prothrombin molecule or, more recently, a few hypo- and dysprothrombinemias by nucleotide sequence analysis of their prothrombin genes. In one example DNA sequence analysis revealed homozygosity for a single basepair (bp) transition in exon 3 at position 1305 (A to G) that predicts the replacement of tyrosine for cysteine at amino acid residue 44 of the mature protein. This mutation in exon 3 of the prothrombin gene is associated with a bleeding tendency and can therefore not be the underlying cause of many unexplained thrombotic events.